Neurturin和NGF对老年性痴呆模型鼠海马突触素表达的影响

为了探讨联合应用 Neurturin和神经生长因子对老年性痴呆模型鼠海马突触素的影响 ,本研究采用切断成年 SD大鼠左侧穹窿海马伞 ,建立隔 -海马胆碱能系统损害的痴呆模型 ,损伤 4周后 ,用免疫组化和图象分析技术等方法对大鼠海马突触素进行定量分析。结果显示 ,损伤对照组损伤侧海马 CA1 区多形层、辐射层、腔隙分子层和齿状回分子层突触素含量分别减少了42 .60 %、46.0 4%、5 7.3 6%和 5 5 .2 2 % ,损伤对照组与正常对照组之间有高度显著性差异 ( P<0 .0 1) ;NGF治疗组损伤侧海马CA1 区多形层、辐射层、腔隙分子层和齿状回分子层突触素含量分别只减少了 3 3 .64 %、3 8.2 5 %、3 8.42 %和 2 7.64 % ,NGF治疗组与损伤对照组之间有显著性差异 ( P<0 .0 5 ) ;联合治疗组损伤侧海马 CA1 区多形层、辐射层、腔隙分子层和齿状回分子层突触素含量分别只减少了 9.97%、8.0 5 %、14 .70 %和 4.2 8% ,联合治疗组与损伤对照组之间有高度显著性差异 ( P<0 .0 1)。结论 :联合应用 neurturin和 NGF促使老年性痴呆模型鼠海马突触素含量增多的效果较单用 NGF治疗好     

神经干细胞移植对192-IgG-saporin老年性痴呆模型鼠的学习记忆及海马胆碱能纤维的影响

目的探讨神经干细胞(NSCs)移植对192-IgG-saporin致老年性痴呆模型鼠学习记忆和海马胆碱能纤维再生的影响。方法采用192-IgG-saporin(2.5μg/5μL)侧脑室注射SD大鼠建立痴呆模型后,行基底前脑神经干细胞移植,4周后行Y迷宫检测,并观察大鼠海马胆碱能纤维数的变化。结果Y迷宫检测显示大鼠的学习、记忆能力,模型组(107.38±9.34、3.75±0.71)与正常组比较明显下降(P<0.01),而移植组(75.26±5.33、5.45±0.51)有所改善(P<0.05);免疫组化显示模型组大鼠海马CA1区辐射层和齿状回分子层胆碱能阳性纤维,模型组与正常组比较,CA1辐射层和齿状回分子层纤维密度分别减少到11.07%和12.96%(P<0.01),与正常组比较均而干细胞移植组则分别恢复到正常组的81.39%和75.30%(P>0.05)。结论神经干细胞能促进192-IgG-saporinAD动物模型鼠学习记忆能力的恢复及海马胆碱能纤维的再生。     

神经干细胞与脑源性神经营养因子联合治疗对老年性痴呆鼠基底前脑一氧化氮合酶阳性神经元的影响

目的探讨神经干细胞（NSCs）与脑源性神经营养因子（BDNF）联合应用对老年性痴呆鼠基底前脑一氧化氮合酶（NOS）阳性神经元的影响.方法切断成年SD大鼠左侧穹窿海马伞（FF）,基底前脑行神经干细胞移植,同时持续侧脑室注射BDNF,4周后行组织化学染色结合图像分析技术观察各组大鼠基底前脑NOS阳性神经元数量和形态学参数的变化.结果损伤后1个月,损伤侧基底前脑内侧隔核（MS）和斜角带垂直支（VDB）内可观察到NOS阳性神经元明显减少,分别为正常组的35.5%和55.8%,（与正常组相比P＜0.01）;移植组NOS阳性神经元数恢复到正常组的74.7%和95.7%,（与损伤组相比P＜0.01）;联合组NOS阳性神经元数达到正常组的115.2%和151.3%,（与移植组相比P＜0.05及P＜0.01）.细胞形态学参数提示,移植组NOS阳性神经元中含部分中等大小的未成熟细胞;联合组中该中等大小的细胞更多.结论神经干细胞移植与BDNF联合治疗,对AD模型鼠基底前脑MS、VDB的NOS阳性神经元有明显的补充和保护作用,并且联合治疗效果比单纯神经干细胞移植效果更佳.     

神经生长因子对痴呆模型鼠海马突触素的影响

切断SD老年鼠(24月龄)左侧穹窿海马伞.造成隔海马胆碱能系统损害的痴呆模型. 用免疫组化和图像分析技术分析神经生长因子对痴呆鼠海马突触素的影响.实验证明损伤一个月后.损伤对照组损伤侧海马CAI区多形层、辐射层、腔隙分子层和齿状回分子层突触素含量分别是减少了28.17% 、32.15%、17.36%和35.22%:NGF治疗组、损伤侧海马CAI区多形层、辐射层、腔隙分子层和齿状回分子层突触素含量只减少了6.17%、5.52%、13.50%和3.81%.提示神经生长因子能够促使痴呆鼠海马内突触素含量的增多.     

BDNF和神经干细胞联用对老年痴呆鼠基底前脑 胆碱能神经元和学习记忆能力的影响

探讨脑源性神经营养因子(BDNF)和神经干细胞(NSCs)联合应用对老年痴呆大鼠基底前脑胆碱能神经元及大鼠学习记忆能力的影响。利用无血清培养技术获得新生SD鼠的海马NSCs,行BrdU标记。切断SD大鼠左侧穹隆海马伞,基底前脑注射NSCs,同时侧脑室注射BDNF,2周后行nestin和BrdU/NF、BrdU/GFAP免疫荧光双标染色,4周后行Y迷宫测试,免疫组化结合图像分析技术观察各组大鼠基底前脑神经营养因子受体(NGFR)阳性神经元数目变化。结果发现,移植后NSCs能够在宿主体内存活且分化成神经元和胶质细胞;免疫组化结果显示损伤组大鼠胆碱能神经元数在内侧隔阂(MS)和斜角带(VDB)分别减少64.3%和49.3%,较正常组明显下降(P<0.01);移植组神经元数下降34.1%和27.6%较损伤组有改善(P<0.05),但与正常组比较有显著性差异(P<0.05);联合组神经元数减少15.1%和18.6%,与正常组无显著性差异(P>0.05)。Y迷宫测试结果显示,大鼠的学习记忆能力与基底前脑NGFR阳性细胞数呈正相关。提示,BDNF和NSCs移植的联用较单独使用NSCs或BDNF更好地改善老年痴呆大鼠的学习记忆能力。     

神经干细胞和BDNF联用对痴呆模型鼠基底前脑小白蛋白表达的影响

探讨神经干细胞(NSCs)和脑源性神经营养因子(BDNF)联用对老年痴呆大鼠基底前脑小白蛋白(PV)表达的影响。利用无血清培养技术获得新生SD大鼠海马NSCs。切断SD大鼠左侧穹隆海马伞,基底前脑注射NSCs,同时侧脑室注射BDNF。4周后行免疫组织化学结合图象分析技术观察各组大鼠基底前脑PV蛋白表达,同时采用半定量RT-PCR法分析基底前脑PVmR-NA表达水平情况。结果发现,损伤组大鼠PV蛋白在内侧隔核和斜角带的表达较正常组明显下降(P<0.01);移植组PV蛋白表达较损伤组有提高(P<0.05),但与正常组比较有显著性差异(P<0.05);联合组PV蛋白表达与正常组无显著性差异(P>0.05)。RT-PCR结果显示,PVmRNA在损伤组和正常组表达量分别为0.2407±0.0825,0.8536±0.1248,两者有显著差异(P<0.01);移植组的表达量为0.5016±0.1009,与损伤组比较差异显著(P<0.05);联合组的表达量较损伤组明显增高(P<0.01),也高于移植组(P<0.05),与正常组比较无显著性差异(P>0.05)。本实验结果提示NSC和BDNF联用较单独使用NSC或BDNF更好地提高PV蛋白及其mRNA的表达,二者具有协同作用。     

神经干细胞移植对注射192-IgG-saporin致AD动物模型鼠基底前脑ChAT阳性神经元的影响

目的探讨神经干细胞(NSCs)移植对注射192-IgG-saporin致老年性痴呆动物模型鼠基底前脑ChAT阳性神经元的影响。方法将24只成年SD大鼠随机分为正常组、模型组和移植组,每组8只。模型组:左侧侧脑室注射免疫毒素192-IgG-saporin2.5μg/5μL,建立192-IgG-saporin致老年性痴呆动物模型;移植组:动物模型建立后,于基底前脑行神经干细胞移植,4周后行免疫组织化学染色,结合图像分析技术观察各组大鼠基底前脑ChAT阳性神经元数量和形态学参数的变化。结果模型组注射192-IgG-saporin后1个月,观察到注射侧基底前脑内侧隔核(MS)和斜角带垂直支(VDB)ChAT阳性神经元明显减少,分别为正常组的9.70%和14.10%(与正常组相比,P<0.01);移植组ChAT阳性神经元恢复到正常组的71.51%和73.52%(与模型组相比,P<0.01)。细胞形态学参数提示,移植组ChAT阳性神经元中含有大小不等的未成熟细胞。结论神经干细胞移植对192-IgG-saporin致痴呆模型鼠基底前脑ChAT阳性神经元有明显补充和保护作用。     

丰富环境干预对孕期手机辐射暴露后仔鼠认知功能的影响

目的:探讨丰富环境干预对孕期长时间手机辐射暴露后仔鼠认知功能的影响。方法:18只SPF级SD孕鼠随机分为两组,分别给予对照处理(不辐射)、长时间手机辐射处理(辐射24 h/d,21d),待仔鼠出生后饲养至18月龄,进行丰富环境干预,共分为3组,分别为对照组(CON)、手机辐射组(PR)及丰富环境组(PR+EE)。于丰富环境干预28 d后应用Morris水迷宫评定仔鼠认知功能;免疫组化及Western Blot检测海马区突触素(SYN)和脑源性神经营养因子(BDNF)表达;透射电镜观察海马区突触的超微结构。结果:与CON组相比,PR组及PR+EE组仔鼠的逃避潜伏期延长,跨越平台的次数减少,SYN和BDNF表达均减少,海马CA1区超微结构改变;而与PR组相比,PR+EE组逃避潜伏期缩短,穿越平台次数增加,SYN和BDNF表达均上调,且SYN阳性细胞增多,海马CA1区突触超微结构改变。结论:丰富环境干预可能通过上调海马SYN和BDNF表达,改变突触超微结构,提高突触可塑性,进而改善孕期手机辐射暴露后仔鼠的认知功能。     

解毒通络方对脑缺血损伤海马区突触可塑性的影响

目的　观察解毒通络方对大鼠脑缺血损伤后海马神经毯的突触形态与突触素P38的表达的影响。方法 :本文应用超微结构分析与免疫组化方法。结果　缺血损伤引起海马CA1区突触前成分溃变、突触数密度下降、突触素P38低水平表达 ;术后 8周 ,模型组与治疗组出现数量较多的新生突触和突触素P38表达水平的提高 ,治疗组突触数密度恢复最为显著 ,已达假手术组的 87 4% ,并明显高于模型组 ;与假手术组比较 ,模型组突触素P38表达COD值提高到 80 % ,而治疗组提高到 114 7% ,治疗组COD值显著高于模型组 (P<0 0 1)。结论　解毒通络方具有促进突触再建和增强、完善再建突触效能的作用     

神经干细胞移植对阿尔茨海默病大鼠海马突触素表达和学习记忆能力的影响

背景:前期实验已证实,移植入阿尔茨海默病大鼠脑内的神经干细胞能够存活、增殖,但其是否可替代损伤或坏死的神经细胞而重建神经通路,改善学习记忆能力尚不清楚。突触素是突触重建的重要标记之一。目的:观察神经干细胞移植对阿尔茨海默病大鼠学习记忆能力及突触表达的影响。方法:SD大鼠随机数字表法分为正常对照组、阿尔茨海默病模型组、2周移植组和4周移植组,除正常对照组外制备阿尔茨海默病模型。另取新生24hSD大鼠海马齿状回分离、培养神经干细胞,经Hoechst33258标记后植入2周和4周移植组海马CA1区,行Y迷宫实验检测大鼠的学习记忆能力,然后取脑进行尼氏染色和突触素免疫组织化学染色。阿尔茨海默病模型组则以同样的方法、同样的位点注入等量无菌生理盐水。正常对照组不施以任何处理。结果与结论:①2周和4周移植组海马CA1区细胞比阿尔茨海默病模型组增多,但仍少于正常对照组(P0.05),平均吸光度与正常对照组相比差异无显著性意义(P0.05)。②2周移植组和4周移植组大鼠海马结构内突触素吸光度值明显高于正常对照组和阿尔茨海默病模型组(P0.05)。③与阿尔茨海默病模型组相比,2周和4周移植组大鼠学习能力和记忆能力均显著增强,正确反应率明显提高(P0.05),而与正常对照组相比,差异无显著性意义(P0.05)。提示移植入脑内的神经干细胞可促进突触形成,改善学习记忆能力。     

穹窿海马伞损伤对大鼠学习记忆和海马GFAP阳性神经胶质细胞的影响

为探讨穹隆海马伞损伤鼠学习记忆能力与海马胶质纤维酸性蛋白阳性细胞之间的关系 ,切断 SD成年大鼠左侧穹窿海马伞 ,用 Y迷宫和免疫组织化学结合图像分析系统测试大鼠学习记忆能力和海马胶质纤维酸性蛋白阳性细胞的变化状况及它们的相互关系。结果显示 :损伤 2周后 ,损伤组损伤侧海马 CA1 区辐射层和齿状回分子层胶质纤维酸性蛋白阳性细胞的数密度较正常组分别增多 3 0 .2 9%和 3 0 .15 % (都为 P<0 .0 1) ,胞体面积分别增加 16.0 4%和 19.42 % (都为 P<0 .0 1) ,齿状回分子层胶质纤维酸性蛋白阳性细胞体密度增大 19.40 % (P<0 .0 5 )。经相关分析 ,大鼠学习记忆能力与海马 CA1 区胶质纤维酸性蛋白阳性细胞数密度呈负相关 (r=-0 .83 6,P<0 .0 1) ,与齿状回数密度呈负相关 (r=-0 .792 ,P<0 .0 1)。提示海马星形胶质细胞可能参与学习记忆过程     

神经生长因子/胶源性神经营养因子基因修饰神经前体细胞单独和联合移植促进阿尔茨海默病模型鼠海马胆碱能纤维网的重建

目的 :观察神经生长因子 (Nervegrowthfactor,NGF) /胶源性神经营养因子 (Glialcellline derivedneurotroph icfactor,GDNF)基因修饰神经前体细胞 (Neuralprogenitorcell,NPC)单独和联合移植对阿尔茨海默病 (Alzheimer’sdisease,AD)模型鼠海马胆碱能纤维网的促重建作用。方法 :将NGF/GDNF基因修饰的NPC单独和联合移植入FF切断的大鼠侧脑室内。移植后三周取海马切片进行AchE纤维组织化学染色。结果 :CA1区和齿状回的NGF组、GDNF组和NGF +GDNF组的AchE纤维数分别为 3 2 %和 5 0 %、1 8%和 2 4 %、2 4 %和 5 8% ,明显高于损伤组(4 %和 6% )和NPC组 (7%和 9% ) ,P均 <0 .0 1 ;NGF组也高于GDNF组 (P <0 .0 5和P <0 .0 1 )。结论 :NGF/GDNF基因修饰的NPC单独和联合移植均能不同程度地促进AD模型鼠海马胆碱能纤维网的重建 ,其中NGF组和NGF +GDNF组作用大于GDNF组     

大鼠背海马内生长抑素mRNA神经元的老年变化

采用原位杂交组织化学及图像定量分析法，研究了大鼠背海马中生长抑素（ＳＳ）ｍＲＮＡ神经细胞的老年变化。年轻大鼠内，ＳＳｍＲＮＡ胞体主要分布于ＣＡ１和ＣＡ２区的锥体层和多形层、ＣＡ３区的辐状层和多形层，以及齿状回的多形层。老年大鼠内，ＳＳｍＲＮＡ胞体则主要集中于背海马的多形层。和年轻大鼠相比，老年大鼠背海马内ＳＳｍＲＡＮ胞体数量显著减少，胞体灰度值明显升高，而胞体截面积无明显年龄变化。结果表明，大鼠背     

GABAergic neurons containing somatostatin-like immunoreactivity in the rat hippocampus and dentate gyrus

Summary The distribution of somatostatin-like immunoreactive (SS-LI) material and its colocalization with glutamic acid decarboxylase (GAD)-like immunoreactivity were studied in the rat hippocampus and dentate gyrus neurons using immunohistochemistry. In the dentate gyrus and CA1 region, SS-LI perikarya were concentrated in the hilus and in the stratum oriens, respectively, whereas immunoreactive cell bodies were rarely seen in other layers. Approximately half of the SS-LI neurons of the CA3 region were situated in the stratum oriens, the other half being scattered in strata pyramidale, lucidum and radiatum. About 90% of SS-LI neurons were also GAD-like immunoreactive, whereas about 14% of GAD-like immunoreactive (GAD-LI) neurons were SS-like immunoreactive. The percentage of GAD-LI neurons which were also immunoreactive for SS varied from one layer to the other. This percentage was about 30% in the hilus of the dentate gyrus and in the stratum oriens of the CA1 and CA3 regions; it was 5–10% in the strata pyramidale, lucidum and radiatum of the CA3 region and reached only 2% in the granule cell layer and molecular layer of the dentate gyrus and in the stratum pyramidale and stratum radiatum in the CA1 region. These observations indicate that the majority of SS-LI neurons in the rat hippocampal formation are a subpopulation of GABAergic neurons.     

BDNF Up-Regulates TrkB Protein and Prevents the Death of CA1 Neurons Following Transient Forebrain Ischemia

The neurotrophin family of growth factors, which includes Nerve Growth Factor (NGF), Brain-Derived Neurotrophic Factor (BDNF), Neurotrophin-3 (NT3) and Neurotrophin-4/5 (NT4/5) bind and activate specific tyrosine kinase (Trk) receptors to promote cell survival and growth of different cell populations. For these reasons, growing attention has been paid to the use of neurotrophins as therapeutic agents in neurodegeneration, and to the regulation of the expression of their specific receptors by the ligands. BDNF expression, as revealed by immunohistochemistry, is found in the pre-subiculum, CA1, CA3, and dentate gyrus of the hippocampus. Strong TrkB immunoreactivity is present in most CA3 neurons but only in scattered neurons of the CA1 area. Weak TrkB immunoreactivity is found in the granule cell layer of the dentate gyrus. Unilateral grafting of BDNF-transfected fibroblasts into the hippocampus resulted in a marked increase in the intensity of the immunoreaction and in the number of TrkB-immunoreactive neurons in the granule cell layer of the dentate gyrus, pre-subiculum and CA1 area in the vicinity of the graft. No similar effects were produced after the injection of control mock-transfected fibroblasts. Delayed cell death in the CA1 area was produced following 5 min of forebrain ischemia in the gerbil. The majority of living cells in the CA1 area at the fourth day were BDNF/TrkB immunoreactive. Unilateral grafting of control mock-transfected or BDNF fibroblasts two days before ischemia resulted in a moderate non-specific protection of TrkB-negative, but not TrkB-positive cells, in the CA1 area of the grafted side. This finding is in line with a vascular and glial reaction, as revealed, by immunohistochemistry using astroglial and microglial cell markers. This astroglial response was higher in the grafted side than in the contralateral side in ischemic gerbils, but no differences were seen between BDNF-producing and non-BDNF-producing grafts. However, grafting of BDNF-producing fibroblasts two days before ischemia significantly and specifically prevented nerve cells from dying in the CA1 area of the ipsilateral hippocampus. Cell survival was associated with increased TrkB immunoreactivity as the majority of living cells were TrkB immunoreactive. Thus, our results show that BDNF is able to up-regulate the expression of TrkB in control and pathological states, and that BDNF prevention of neuronal death following transient forebrain ischemia is associated with increased expression of its specific receptor.     

NOV蛋白在切割海马伞大鼠海马中表达的变化

通过切割右侧海马伞制备大鼠海马伞损伤模型,应用Westernblotting、免疫组织化学技术,观察切割海马伞后不同时程海马中肾母细胞瘤过度表达基因(NOV)蛋白表达的变化,并对结果进行图像处理和统计学分析。Westernblotting结果显示,NOV蛋白在切割海马伞后3d表达开始上升,14d达高峰后缓慢下降。切割海马伞后14d切割侧和正常侧相比较,海马CA1-CA3区的锥体细胞层及齿状回颗粒层NOV阳性细胞数目无显著性差异(P>0.05),但切割侧NOV阳性细胞明显比正常侧深染,两侧比较平均灰度值有显著性差异(P<0.01)。结合本课题组以往的工作,本研究结果提示,切割海马伞后海马中高表达的NOV蛋白可能参与了诱导神经干细胞迁移和向神经元分化的过程。     

复方丹参对脑缺血再灌注后大鼠海马和齿状回神经细胞凋亡及Bcl-2 mRNA表达的影响

目的:探讨中药复方丹参对大鼠脑缺血再灌注后海马和齿状回神经细胞凋亡及Bcl-2 mRNA表达的影响。方法:采用大脑中动脉内栓线法建立大鼠大脑中动脉缺血再灌注模型,应用原位细胞凋亡检测和原位杂交技术检测大鼠海马和齿状回神经细胞凋亡和Bcl-2 mRNA的表达并做图像分析。结果:与假手术对照组比较,缺血再灌注组凋亡神经细胞主要位于缺血侧海马CA1、CA3区,齿状回凋亡细胞较少。3个区神经细胞Bcl-2mRNA的表达在缺血再灌注2 h后升高,随时间的延长逐渐增强。复方丹参组神经细胞Bcl-2 mRNA的表达明显强于缺血再灌组,而凋亡神经细胞数明显较低。结论:复方丹参可通过上调神经细胞Bcl-2 mRNA的表达,抑制神经细胞凋亡,从而减轻缺血再灌注对大鼠海马和齿状回的损伤。     

Brain-derived neurotrophic factor, nerve growth and neurotrophin-3 selected regions of the rat brain following kainic acid-induced seizure activity.

Changes in levels of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF) and neurotrophin-3 (NT-3) in various regions of the rat brain following kainic acid-induced seizure activity were investigated. BDNF protein, as measured by a two-site enzyme immunoassay, increased transiently 12-24 h after the intraperitoneal administration of kainic acid to 61.6 ng/g wet weight in the hippocampus (approximately 10-fold increase), 19.5 ng/g in the piriform plus entorhinal cortex (approximately 10-fold) and 8.2 ng/g in the olfactory bulb (approximately 16-fold), and then rapidly decreased. Increases of 2- to 4-fold in levels of BDNF were also detected in the septum, cerebral cortex, striatum and hypothalamus, but not in the cerebellum. In contrast, levels of NGF and NT-3 decreased 24 h after the administration of kainic acid. Western and Northern blotting analyses of hippocampal tissues, respectively, revealed increase in levels of a 14-kDa protein corresponding to BDNF and its mRNA at both 4.2 and 1.4 kb. Hippocampal mRNAs for NGF and NT-3 increased and decreased, respectively, in kainic acid-treated rats. Immunohistological investigations showed that, in the hippocampus, the administration of kainic acid enhanced a homogeneous immunoreactivity of BDNF in the polymorph inner layer (the stratum radiatum of the CA3/CA4 regions and the hilar region) and in granule cells of the dentate gyrus. BDNF protein was found in neurons, but not at all in glial cells or in blood vessels, and was localized in the cytoplasm, the nucleoplasm and the primary dendrites of neurons as well as in perisynaptic extracellular spaces, but hardly in their axons. Our results show that kainic acid treatment increases levels of BDNF, but not NGF or NT-3, in various regions of the rat brain, other than the cerebellum. Also, the majority of BDNF newly synthesized by hippocampal granule neurons is secreted into the perisynaptic extracellular space in the polymorph inner layer of the dentate gyrus, supporting an autocrine-like role for the factor in synaptic functions.